Kidney Organoid Modeling of WT1 Mutations Reveals Key Regulatory Paths Underlying Podocyte Development.

Wilms tumor-1(WT1) is a crucial transcription factor that regulates podocyte development. However, the epigenomic mechanism underlying the function of WT1 during podocyte development has yet to be fully elucidated. Here, single-cell chromatin accessibility and gene expression maps of foetal kidneys and kidney organoids are generated. Functional implications of WT1-targeted genes, which are crucial for the development of podocytes and the maintenance of their structure, including BMPER/PAX2/MAGI2 that regulates WNT signaling pathway, MYH9 that maintains actin filament organization and NPHS1 that modulates cell junction assembly are identified. To further illustrate the functional importance of WT1-mediated transcriptional regulation during podocyte development, cultured and implanted patient-derived kidney organoids derived from the Induced Pluripotent Stem Cell (iPSCs) of a patient with a heterozygous missense mutation in WT1 are generated. Results from single-cell RNA sequencing (scRNA-seq) and functional assays confirm that the WT1 mutation leads to delays in podocyte development and causes damage to cell structures, due to its failure to activate the targeting genes MAGI2, MYH9, and NPHS1. Notably, correcting the mutation in the patient iPSCs using CRISPR-Cas9 gene editing rescues the podocyte phenotype. Collectively, this work elucidates the WT1-related epigenomic landscape with respect to human podocyte development and identifies the disease-causing role of a WT1 mutation.

Clinical Implications of a New DDX58 Pathogenic Variant That Causes Lupus Nephritis due to RIG-I Hyperactivation.

BACKGROUND: Lupus nephritis (LN) is one of the most severe complications of systemic lupus erythematosus, with heterogeneous phenotypes and different responses to therapy. Identifying genetic causes of LN can facilitate more individual treatment strategies. METHODS: We performed whole-exome sequencing in a cohort of Chinese patients with LN and identified variants of a disease-causing gene. Extensive biochemical, immunologic, and functional analyses assessed the effect of the variant on type I IFN signaling. We further investigated the effectiveness of targeted therapy using single-cell RNA sequencing. RESULTS: We identified a novel DDX58 pathogenic variant, R109C, in five unrelated families with LN. The DDX58 R109C variant is a gain-of-function mutation, elevating type I IFN signaling due to reduced autoinhibition, which leads to RIG-I hyperactivation, increased RIG-I K63 ubiquitination, and MAVS aggregation. Transcriptome analysis revealed an increased IFN signature in patient monocytes. Initiation of JAK inhibitor therapy (baricitinib 2 mg/d) effectively suppressed the IFN signal in one patient. CONCLUSIONS: A novel DDX58 R109C variant that can cause LN connects IFNopathy and LN, suggesting targeted therapy on the basis of pathogenicity. PODCAST: This article contains a podcast at.

Generation of induced pluripotent stem cell line (NCKDi001-A) from a 19-year-old patient with a novel COL4A5 gene mutation in Alport syndrome.

The clinical manifestations of Alport syndrome may vary depending on the involved organs such as the kidneys, cochlea and eyes. The pathogenic genes involved are those encoding different chains of type IV collagen. We collected PBMCs of a patient with a novel COL4A5 gene mutation(c.2687G > C). Subsequently, we used the electroporation system to transfer the reprogramming plasmids expressing OCT3/4, SOX2, KLF4, LIN28 and L-MYC into the PBMCs. We simultaneously carried out the tests on the iPSCs including Sanger sequencing for confirming the mutation site, immunofluorescence assay and flow cytometry for pluripotency markers as well as teratoma experiment for validating the pluripotency.

Nocardiosis in patients with nephrotic syndrome: a retrospective analysis of 11 cases and a literature review.

OBJECTIVES: We evaluated the clinical manifestations and outcomes of nocardiosis, a rare opportunistic infection that occurs in patients with nephrotic syndrome. METHODS: The records of NS patients with nocardiosis in a single hospital during 2000-2019 were retrieved and studied in detail. RESULTS: Eleven patients were included. The mean time to develop nocardiosis after glucocorticoid therapy was 11.5 ± 14.8 months. Most patients had fever, elevated white blood cell counts and C-reactive protein, whereas procalcitonin levels were normal or slightly elevated in 91% (10/11) patients, except one patient suffered from septic shock. Nine patients were tested for CD4+ T-cell counts; of these, four patients had counts < 200 cells/µL. The most common site of nocardiosis involvement was lung (100%), followed by subcutaneous tissue (72.7%). Radiological findings for lungs in seven cases were characterized by isolated or scattered nodules and masses, usually located subpleural or close to the hilum. Positive smears of Nocardia were detected in 100% of samples of subcutaneous abscess and pleural fluid. Nine patients received oral trimethoprim-sulfamethoxazole, four of which received combined carbapenem, and the remaining two patients received carbapenem monotherapy. The long-term prognosis was excellent, with a treatment success rate of 100% in all patients. CONCLUSIONS: NS patients can develop immunodeficiency after treatment with glucocorticoid and immunosuppressants. In cases where patients develop systemic multiple abscesses, or lung images reveal isolated or scattered nodules and masses that are subpleural or close to the hilum, nocardial infection should be considered. Early diagnosis and specific treatment may improve patient outcomes.

A Novel COL4A5 Splicing Mutation Causes Skipping of Exon 14 in a Chinese Family with Alport Syndrome.

BACKGROUND: Alport syndrome (AS) is an inherited progressive renal disease caused by mutations in COL4A3, COL4A4, and COL4A5. Although mutation screening in the genes responsible for AS is typically performed, only a small proportion of patients receive genetic testing in China, and the functional consequences of multiple splicing variants in AS patients have not been investigated. METHODS: A family with X-linked AS was diagnosed based on family history and pathological findings from a kidney biopsy. Targeted next-generation sequencing was used to identify the causative mutation, and a minigene assay was performed to test the influence of the mutation on splicing. RESULTS: A c.834+2T>G in COL4A5 was identified and shown to co-segregate with AS in the family. The variant is located in the canonical splicing site and is predicted to induce aberrant splicing. Minigene assay using HEK 293T cells indicated the skipping of exon 14 in -COL4A5. CONCLUSIONS: The novel COL4A5 splicing mutation identified in the current study broadened the genetic spectrum of X-linked AS and further deepened our insight of the disease's molecular mechanism.

Autosomal Dominant Tubulointerstitial Kidney Disease Due to UMOD Mutation: A Two-Case Report and Literature Review.

Autosomal dominant tubulointerstitial kidney disease due to UMOD (encoding uromodulin) mutation (ADTKD-UMOD) is a rare hereditary disease. In the present study, we reported 2 ADTKD cases with confirmed UMOD mutations (Arg185His, Trp258Gly) by gene testing. They were young men and presented with hyperuricemia and renal dysfunction with no hematuria or proteinuria. Renal histology showed chronic tubulointerstitial nephropathy with fibrillar inclusions in the cells of distal tubules. Electron microscopy illustrated extensive bundled and cystic endoplasmic reticulum. Immunohistological analysis confirmed intracytoplasmic aggregates of uromodulin in the distal tubules. Since ADTKD-UMOD is an underdiagnosed disease, electron microscopy and immunohistochemical staining for uromodulin are helpful in the diagnosis of ADTKD-UMOD and genetic analysis is the gold standard.

A Case of Switch from C3 Glomerulonephritis to Proliferative Glomerulonephritis with Monoclonal IgG Deposits.

Monoclonal immunoglobulins have been implicated in the development of C3 glomerulonephritis (C3GN) and Proliferative glomerulonephritis with monoclonal IgG deposits (PGNMID). We report a 58-year-old female who showed a switch from C3GN to PGNMID. She presented with mild proteinuria and normal renal function for the first time. Her renal biopsy showed a severe mesangial proliferation with isolated C3 deposits, thus being diagnosed as C3GN. Two years later, her condition became serious. Repeat renal biopsy showed a membranoproliferative glomerulonephritis with deposition of the κ light chain of IgG3 in the glomeruli. She was diagnosed with proliferative glomerulonephritis with monoclonal IgG deposits (IgG3-κ). This case demonstrates that there are several types of monoclonal gammopathy (MGP)-associated glomerulonephritis, and they can switch among each other in some patients.

Risk Factors for Renal Survival in Chinese Patients with Myeloperoxidase-ANCA-Associated GN.

BACKGROUND AND OBJECTIVES: Our study explored the association of histopathologic classification of ANCA-associated GN with renal survival in Chinese patients with myeloperoxidase-ANCA-associated GN. DESIGN, SETTING, PARTICIPANTS, & MEASUREMENTS: Two hundred fifteen patients with biopsy-proven myeloperoxidase-ANCA-associated GN were included from January of 1996 to December of 2014. The biopsies included focal (n=27), mixed (n=82), crescentic (n=47), and sclerotic (n=59) classes. The long-term renal outcome and risk factors of myeloperoxidase-ANCA-associated GN for different histopathologic classes were retrospectively analyzed. RESULTS: During a median follow-up time of 22 (9-51) months, 88 (40.9%) patients reached ESRD. The 5-year renal survival (overall 58.7%) was highest in the focal class (100.0%) and lowest in the sclerotic class (20.7%), with no difference between the mixed (58.9%) and crescentic (67.4%) classes. Patients in the mixed (hazard ratio, 0.34; 95% confidence interval, 0.20 to 0.57; P<0.001) and crescentic (hazard ratio, 0.31; 95% confidence interval, 0.16 to 0.59; P<0.001) classes were at lower risk for ESRD compared with patients in the sclerotic class, as were patients who received glucocorticoids plus mycophenolate mofetil (hazard ratio, 0.32; 95% confidence interval, 0.18 to 0.60; P<0.001) compared with those receiving glucocorticoids alone. In addition, patients with a serum creatinine level ≥4 mg/dl (hazard ratio, 2.93; 95% confidence interval, 1.77 to 4.85; P<0.001) or hypoalbuminemia (hazard ratio, 2.11; 95% confidence interval, 1.32 to 3.34; P=0.002) were at higher risk for ESRD. A serum creatinine level ≥4 mg/dl and a percentage of global sclerotic glomeruli ≥60% were the two independent risk factors for ESRD in the sclerotic class. CONCLUSIONS: The histopathologic classification of ANCA-associated GN in combination with serum creatinine and serum albumin levels and treatment regimen is associated with renal outcome in myeloperoxidase-ANCA-associated GN. The evaluation of serum creatinine level and percentage of global sclerotic glomeruli provides additional information on the risk of renal survival in the sclerotic class of myeloperoxidase-ANCA-associated GN.